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Kaili Wu, Zhongting Li, Fang Duan, Jingyu Liao, Qiang Huang; Effective delivery of small interfering RNA to mouse cornea by eye drops. Invest. Ophthalmol. Vis. Sci. 2013;54(15):2725.
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The transfection of siRNA in vivo is essential for gene therapy. However, the successful delivery of siRNA in ocular surface is still very difficult to achieve. In the current study, we sought to investigate the efficacy of deliver small interference RNA (siRNA) to the mouse cornea epithelium by eye drops that contain siRNA and different cationic complexing agents.
The mouse (BALB/c) corneal epithelial layer was mechanical damaged in 2 mm diameter in the center. After 12 hours, eye drops, including 20μmol/l cy3 labeled siRNA (cy3-siRNA) with different transfection reagents (Lipofectamine 2000, EntransterTM-in vivo, PEI, in vivo-jetPEI and PEO-PPO-PEO polymeric micelles), were applied for various times (4 times a day for two days and 8 times a day for one or two days). The transfection efficiency was examined by fluorescence microscopy of the corneal flat mount and frozen section. In addition, the transfection of the infected-cell polypeptide 4-targeting small interfering RNA (ICP4-siRNA) in mouse herpes simplex keratitis (HSK) was examined using the above method. The antiviral effects were evaluated by in vivo obsevation, microscopy examination and reverse transcription polymerase chain reaction (RT-PCR).
Compared with the other transfection reagents, 0.75mg/ml PEI with cy3-siRNA complex that was instilled 8 times daily for two days revealed the highest fluorescence in cornea epithelial cells (p<0.01). In mouse HSK, application of ICP4-siRNA+PEI eye drops reduced the damage of cornea epithelia, leading to recovery of cornea ulcer, and decreased the viral VP16 expression in cornea tissue (P < 0.05). Meanwhile, we also found that cornea became cloudy in the eye instilled with PEI.
PEI can effectively deliver the siRNA into the cornea epithelium by eye drops. ICP4-siRNA transfection can inhibit HSV-1 replication in vivo
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